It is known that peptide aldehydes of specific structure have the ability to inhibit the proteolytic reactions of serine or cysteine enzymes. The mechanism of enzyme inhibition is assumed to be an addition reaction between the reactive OH or SH group of the enzyme and the -CH.dbd.O group of peptide aldehydes, the hemiacetal formed upon addition being the "non-productive" analogue of the tetrahedral transitory complex formed in the course of enzyme-substrate interaction (Westernik and Wolfenden: J. Biol. Chem. 247, 8195 (1972)). The first members of this family of compounds were the leupeptines of natural origin, i.e. acetyl- and propionyl-L-leucyl-L-leucyl-arginine aldehyde hydrochlorides, which were both capable to inhibit plasmin, trypsin and papain (Kawamura et al.: Chem. Pharm. Bull. 17, 1902 (1969); H. Umezawa: Enzyme Inhibitors of Microbial Origin, University Park Press, Baltimore-London-Tokyo, 1972, pp. 17 to 29). Other peptidyl-arginine aldehydes, e.g. benzoyl-D-allo-isoleucyl-L-prolyl-L-arginine aldehyde p-toluenesulfonate and D-phenylalanyl-L-prolyl-L-arginine aldehyde acetate, exhibit marked antithrombin activity (Hungarian Patent No. 169,870).
On the basis of the NMR studies of leupeptines it was assumed that the acyl-arginine aldehyde hydrochlorides are mixtures of varying composition (Maeda et al.: J. Antibiotics (Tokyo), 24, 402/1971/; Kawamura et al.: Chem. Pharm. Bull., 17, 1902 (1969). In addition to the acyl-arginine aldehyde hydrochloride there are two other components present, the aldehyde hydrate as well as the cyclic carbinolamine. As regards these components the aldehyde hydrate cannot directly participate in hemiacetal formation leading to enzyme inhibition (Westernik and Wolfenden: J. Biol. Chem. 247, 8195/1972/). This ever varying rate and grade of aldehyde hydrate formation may be the cause of the recently observed phenomenon that both the enzyme inhibitory activity of the above synthetic compounds and the antithrombin effect of D-phenylalanyl-L-prolyl-L-arginine aldehyde acetate or hydrochloride for instance, are changing, and that their potency is reduced upon standing in solution [tris-(hydroxy-methyl)-methylamine hydrochloride, i.e. TRIS/HCl buffer, pH=7.4].